New analytical methodologies involving immobilized enzyme reactors (IMER) and high performance liquid chromatography (HPLC) are being explored for use in biomedical analysis and clinical chemistry. As pointed out previously, this combination could be synergistic since the separation step would remove interferences which might be present in the sample. Use of class specific enzyme reactions will allow specific detection reactions, decreasing the necessity of pre-chromatographic clean-up or complex separation systems. The immobilized enzyme systems will be studied in mixed aqueous-organic solvent systems in the hope of elucidating some enzyme structure-function relationships. Specific projects undertaken are the use of 3 alpha-hydroxysteroid dehydrogenase as a post-column reaction for detection of bile acids and development of an immobilized beta-glucuronidase system for hydrolysis of steroid glucuronides in urine prior to analysis.